Thermostable bacterial L-asparaginase for polyacrylamide inhibition and in silico mutational analysis.

Abstract

The L-asparaginase (ASPN) enzyme has received recognition in various applications including acrylamide degradation in the food industry. The synthesis and application of thermostable ASPN enzymes is required for its use in the food sector, where thermostable enzymes can withstand high temperatures. To achieve this goal, the bacterium Bacillus subtilis was isolated from the hot springs of Tapovan for screening the production of thermostable ASPN enzyme. Thus, ASPN with a maximal specific enzymatic activity of 0.896 U/mg and a molecular weight of 66 kDa was produced from the isolated bacteria. The kinetic study of the enzyme yielded a Km value of 1.579 mM and a Vmax of 5.009 µM/min with thermostability up to 100 min at 75 °C. This may have had a positive indication for employing the enzyme to stop polyacrylamide from being produced. The current study has also been extended to investigate the interaction of native and mutated ASPN enzymes with acrylamide. This concluded that the M₁₀ (with 10 mutations) has the highest protein and thermal stability compared to the wild-type ASPN protein sequence. Therefore, in comparison to a normal ASPN and all other mutant ASPNs, M₁₀ is the most favorable mutation. This research has also demonstrated the usage of ASPN in food industrial applications.